DESCRIPTION: The proposed research seeks to describe the molecular phenotype of single cells in cell populations (e.g., organs and tissues) through the direct measurement of the lipidome and metabolome at cellular spatial resolution using MALDI mass spectrometry. Recent innovations in instrumentation conceived and built at Vanderbilt University (VU) uniquely provide the sensitivity, spatial resolution, and throughput required to meet this analytical challenge. We have developed two instrumental approaches that enable single cell analysis by MS. First, advanced laser optical systems developed in our laboratory permit targeting and ablation of features as small as a single cell with subsequent MS of the analytes using a time-of-flight mass spectrometer. This technology has sufficient spatial resolution to target single human cells in tissue without microdissection; however, sensitivity is limited due to the minimal amount of ablated material. Second, to address the need for greater sensitivity, we will employ a technique called continuous accumulation of selected ions (CASI) in conjunction with an FT-ICR mass analyzer. CASI utilizes gas-phase ion enrichment where a single mass-to- charge (m/z) range is isolated and stored in a linear ion trap prior to detection, increasing sensitivity up to 1000-fold and providing the sensitivity to measure a variety of analytes from single cells. We propose to integrate these two instrumental advances on a single instrument platform and will validate this novel platform specifically for the analysis of single cells isolated from in pancreatic islets and through the direct analysis of single cell directly from pancreas tissue sections.